The present invention relates to isolated antibodies specifically binding to human HER3, methods for performing immunohistochemistry using these antibodies, and methods for their production.
Human HER3 (ErbB-3, ERBB3, c-erbB-3, c-erbB3, receptor tyrosine-protein kinase erbB-3, SEQ ID NO: 17) encodes a member of the epidermal growth factor receptor (EGFR) family of receptor tyrosine kinases which also includes HER1 (also known as EGFR), HER2, and HER4 (Kraus, M. H. et al., PNAS 86 (1989) 9193-9197; Plowman, G. D. et al., PNAS 87 (1990) 4905-4909; Kraus, M. H. et al., PNAS 90 (1993) 2900-2904). Like the prototypical epidermal growth factor receptor, the transmembrane receptor HER3 consists of an extracellular ligand-binding domain (ECD), a dimerization domain within the ECD, a transmembrane domain, an intracellular protein tyrosine kinase domain (TKD) and a C-terminal phosphorylation domain. This trans-membrane protein has a Heregulin (HRG) binding domain within the extracellular domain but not an active kinase domain. It therefore can bind this ligand but not convey the signal into the cell through protein phosphorylation. However, it does form heterodimers with other HER family members which do have kinase activity. Heterodimerization leads to the activation of the receptor-mediated signaling pathway and transphosphorylation of its intracellular domain. Dimer formation between HER family members expands the signaling potential of HER3 and is a means not only for signal diversification but also signal amplification. For example the HER2/HER3 heterodimer induces one of the most important mitogenic signals via the PI3K and AKT pathway among HER family members (Sliwkowski M. X., et al., J. Biol. Chem. 269 (1994) 14661-14665; Alimandi M, et al., Oncogene. 10 (1995) 1813-1821; Hellyer, N. J., J. Biol. Chem. 276 (2001) 42153-4261; Singer, E., J. Biol. Chem. 276 (2001) 44266-44274; Schaefer, K. L., Neoplasia 8 (2006) 613-622).
Amplification of this gene and/or overexpression of its protein have been reported in numerous cancers, including prostate, bladder, and breast tumors. Alternate transcriptional splice variants encoding different isoforms have been characterized.
One isoform lacks the intermembrane region and is secreted outside the cell. This form acts to modulate the activity of the membrane-bound form. Additional splice variants have also been reported, but they have not been thoroughly characterized.
Despite the fact that human HER3 is known since more than twenty years it has been and even today is extremely difficult to detect the protein HER3 in a tissue sample. However, detecting HER3 in a tissues sample is crucial to correlate structure and morphology of a given sample with the localization, tissue distribution and/or concentration of HER3 to a biological function, in particular to a pathophysiological context related to numerous cancers.
While several anti-HER3 antibodies are available as research reagents from various companies, no satisfactory staining of a tissue sample, especially of a tissue sample that has been formalin-fixed and paraffin-embedded would appear to be possible using those reagents. In particular there is a need for antibodies showing a high binding specificity and sensitivity towards HER3 when used in automated staining systems for tissue samples such as Ventana Benchmark XT.
It was the task of the present invention to identify an antibody that specifically binds to the human HER3 protein and that can at least partially overcome the problems known in the art.